Chimeric cDNA which contain portions of both wild-type and attenuated HAV cDNA were constructed. RNA transcripts from several of these cDNA constructs were infectious when transfected into monkey kidney cells and the results indicated that two genes, 2B and 2C, were important for tissue culture adaptation. Site- directed mutagenesis and transfection assays suggested that at most, four of six base changes in the attenuated clone are sufficient to allow wild-type virus to grow in cell culture. These mutations will be inserted by site-directed mutagenesis into the wild-type clone and transfection assays will be done to determine the minimum number of mutations which allow growth in tissue culture. Viruses from the transfection experiments will be inoculated into marmosets to determine which portions of the HAV genome are associated with attenuation and how this phenotype relates to the ability to grow in cell culture. The 2B and 2C genes of the cell adapted HAV will be used to permanently transform a continuous monkey cell line in an attempt to generate a complementing cell line which supports the growth of wild-type virus or improves the growth of the cell adapted virus.